Abstract Book 2010 - CIMT Annual Meeting

092 Tomsitz | Tumor biology & interaction with the immune system
Establishment of a pre-clinical NOD/LtSz-scid IL2Rgammac null
(NSG) mouse model to evaluate immunotherapeutic strategies
for acute myeloid leukemia (AML)
Dirk Tomsitz 1 , Ariane Brunk 1 , Marion Nonn 1 , Alexander Hohberger 1 , Shamsul A. Khan 1 ,
Eva Distler 1 , Matthias Theobald 1 , Wolfgang Herr 1 , Udo F. Hartwig 1
1 Dept. of Medicine III - Hematology and Oncology, Johannes-Gutenberg-University Medical Center,
Mainz, Germany
Introduction: Acute myeloid leukemia (AML) is a
cancer of the myeloid line of blood cells, characte-
rized by the clonal expansion of immature mye-
loblasts potentially initiating from rare leukemic
stem cells. Xenotransplantation of human AML into
immunodeficient mice is essential for establishing
a preclinical model to i) assess the potency of modified
donor lymphocyte grafts to induce immunotherapeutic
graft-versus-leukemia (GVL)-reactivity
and ii) to study the properties of leukemic stem
cell biology. Here we report the successful engraftment
of 8 out of 14 patient-derived, primary AML
samples in NOD/LtSz-scid IL2Rgammacnull (NSG)
mice within 2-12 weeks, with a mean of 0,2-40%
human leukemic cells in the bone marrow (BM). In
first studies, we also show the successful eradication
of engrafted AML-blasts using AML-reactive,
HLA-mismatched T lymphocytes.
Methods: Mice were used unconditioned or irradiated
with 150 cGy 4-16h prior to intravenous (i.v.)
transfer of 5x104-1x107 primary AML cells. Engraftment
kinetics and dose-dependencies were analyzed
by means of clinicopathological criteria and
flow cytometry. AML-reactive T cells were generated
in a HLA-mismatched setting over 3 weeks from
PBMCs isolated from buffy coats. One week after
transplantation of 5x105 AML cells from patient
MZ667, AML-bearing mice were i.v. injected with
5x106 CD8+ leukemia-reactive T lymphocytes.
Results: Stable AML-engraftment in NSG recipients
was detected upon transplantation of 8 out of 14
primary AML samples by determing the percentage
of CD33+/CD45+ cells in blood, spleen and BM.
Expression of Flt3 mutations appeared to promote
engraftment of AML-samples. In addition to leukemic
blasts, we frequently observed co-engraftment
of human T cells present in the AML-graft, possib-
ly facilitated due to irradiation of recipients. In 4
cases, stable AML engraftment was inhibited by
T cell outgrowth and induction of xenoreactivity.
However, in general irradiation augmented the
engraftment efficacy of AML. Thus, we attempted
to circumvent T cell growth by a) depleting T cells
from AML-samples using MACS-technology® or b)
treating animals with immunosuppressiva such as
Cyclosporin A, Mycophenolatemofetil and Tacrolimus.
Immunomagnetic T cell depletion resulted in
stable but decelerated AML engraftment without
T cell outgrowth, whereas preliminary results indicated
that treatment of transplanted mice with
immunsuppressiva abrogate engraftment of both
AML and T cells. Finally, we started to explore our
model for investigating GVL-responses of AMLreactive
human T cell lines generated by repetitive
stimulation of healthy donor lymphocytes with
AML blasts. In first studies using HLA-mismatched
AML MZ667-reactive CD8+ T cells we could demonstrate
that following adoptive transfer of T
lymphocytes into AML MZ667 bearing mice, leukemic
blasts could be completely eradicated, whereas
in controls stable engraftment was detectable in the
BM.
Conclusions: These results suggest that NSG mice
represent a valuable tool for the establishment of a
preclinical AML model to evaluate the immunotherapeutic
GVL-reactivity of ex vivo modified T cellgrafts
as well as to analyze properties of leukemia
stem cell biology.
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